Process for oxidation of steroidal compounds having allylic groups

ABSTRACT

The instant invention involves a process for oxidizing compounds containing an allylic group, i.e. those containing an allylic hydrogen or allylic alcohol group, to the corresponding enones, using a ruthenium-based catalyst in the presence of a hydroperoxide. Particularly, Δ-5-steroidal alkenes can be oxidized to the corresponding Δ-5-7-keto alkenes.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application is a national phase application under 35 U.S.C.§371 of PCT application PCT/US95/06004, filed May 15, 1995, which inturn claims priority to continuation U.S. patent application Ser. No.08/245,935, filed May 19, 1994, now abandoned.

FIELD OF THE INVENTION

The present invention is concerned with a novel process for thecatalytic oxidation of compounds containing an allylic group usingruthenium based catalysts. The process is generally useful for theoxidation of compounds containing allylic hydrogens or alcohols, andparticularly for Δ-5 steroidal compounds.

BACKGROUND OF THE INVENTION

The principal mediator of androgenic activity in some target organs,e.g. the prostate, is 5α-dihydrotestosterone (“DHT”), formed locally inthe target organ by the action of 5α-reductase, which convertstestosterone to DHT. Certain undesirable physiological manifestations,such as acne vulgaris, seborrhea, female hirsutism, androgenic alopeciawhich includes female and male pattern baldness, and benign prostatichyperplasia, are the result of hyperandrogenic stimulation caused by anexcessive accumulation of testosterone (“T”) or similar androgenichormones in the metabolic system. Inhibitors of 5α-reductase will serveto prevent or lessen symptoms of hyperandrogenic stimulation in theseorgans. See especially U.S. Pat. No. 4,377,584, issued Mar. 22, 1983,and U.S. Pat. No. 4,760,071, issued Jul. 26, 1988, both assigned toMerck & Co., Inc. It is now known that a second 5α-reductase isozymeexists, which interacts with skin tissues, especially in scalp tissues.See, e.g., G. Harris, et al., Proc. Natl. Acad. Sci. USA, Vol. 89, pp.10787-10791 (November 1992). The isozyme that principally interacts inskin tissues is conventionally designated as 5α-reductase 1 (or5α-reductase type 1), while the isozyme that principally interactswithin the prostatic tissues is designated as 5α-reductase 2 (or5α-reductase type 2).

The oxidation of Δ-5-steroidal alkenes to the corresponding enones is animportant step in the synthesis of steroid end-products useful as5α-reductase inhibitors. Chromium based oxidations have previously beenused for the oxidation of allylic groups, but are environmentallyunacceptable and require silica gel chromatography. The instantinvention provides an improved alternative method for oxidizingΔ-5-steroidal alkenes, which is convenient to run, and isenvironmentally friendly. Furthermore, the yield and purity of theoxidized intermediate obtained by the instant process meets or exceedsthose obtained when other previously known oxidation methods are used.

SUMMARY OF THE INVENTION

The novel process of this invention involves the oxidation of compoundscontaining an allylic alcohol group or allylic hydrogens to thecorresponding enones using a ruthenium based catalyst in the presence ofa hydroperoxide. Particularly, this invention involves conversion ofΔ-5-steroidal alkenes to Δ-5-7-keto-steroidal alkenes, using a rutheniumbased catalyst in the presence of a hydroperoxide. This novel processcan be exemplified in the following embodiment:

Compounds of Formula II are useful as intermediates in the preparationof 7β-substituted 3-keto-4-azasteroid compounds, such as those which are5α-reductase inhibitors. 5α-Reductase inhibitors are useful in thetreatment of hyperandrogenic disorders such as benign prostatichyperplasia, acne vulgaris, seborrhea, female hirsutism, androgenicalopecia, male pattern baldness, and the prevention and treatment ofprostatic carcinoma.

DETAILED DESCRIPTION OF THE INVENTION

The novel process of this invention involves the discovery thatsteroidal compounds containing a C5-C6 double bond (i.e., Δ-5-steriodalalkenes) can be oxidized to the corresponding 7-keto compounds bytreatment with a hydroperoxide in the presence of a ruthenium-basedcatalyst. Using the same process, compounds containing an allylicalcohol group can likewise be oxidized to their corresponding ketones.For reference, the standard numbering around the unsubstituted coresteroid structure and the letter designation of the rings is as follows:

It has surprisingly been discovered that the instant oxidation processwill proceed using any catalyst which is ruthenium based. Many rutheniumbased catalysts are known in the art, and any such ruthenium basedcatalyst can be used with the instant process. Examples of rutheniumbased catalysts that may be used in this process include but are notlimited to the following: RuW₁₁O₃₉SiNa₅, RuCl₃, RuCl₂(PPh₃)₃, Ru(acac)₃,Ru(dimethylglyoximato)₂(PPh₃)₂, RuO₂, Ru(TPP)(CO)(THF), Ru(bipy)₂Cl₂,Ru(TPP)(CO)(THF), Ru/C and K₅SiRu(H₂O)W₁₁O₃₉. “TPP” istetraphenylporphine; “acac” is acetylacetonate; “bipy” is bipyridine.Ruthenium based catalysts are described in, e.g., R. Neuman, J. Am.Chem. Soc., Vol. 112, 6025 (1990); S-I. Murahashi, Tetrahedron Letters,Vol. 34, 1299 (1993).

Particularly, a ruthenium sodium tungstate-based catalyst is used, andmore particularly RuW₁₁O₃₉SiNa₅. A catalytic amount of the rutheniumcompound is used in this reaction. Those skilled in the art are familiarwith the use of catalytic amounts of reaction catalysts, and willappreciate that the amount of catalyst that can be used may vary withthe scale of the reaction and the particular ruthenium based catalystemployed. An exemplary amount of the ruthenium based catalyst rangesfrom about 0.05 to 5 mol %, and particularly about 0.5 mol % of catalystper mole % of starting material, but variations beyond this range wouldbe acceptable as well.

The alkene starting material is treated with a hydroperoxide in thepresence of the ruthenium-based catalyst for conversion to thecorresponding enone. Many hydroperoxides are known in the art, and anysuch hydroperoxide can be used with the instant process. Examples ofhydroperoxides that may be used in this process include but are notlimited to t-butyl hydrogen peroxide (t-BuOOH), cumene hydroperoxide,hydrogen peroxide, and benzoyl peroxide, with t-BuOOH being preferred.An amount of hydroperoxide sufficient to complete the oxidation shouldbe used, for example at least about 2 moles, and preferably about 8 to10 moles per mole of starting material.

Any commercially available solvent or combinations thereof may beemployed in the instant process step, such as alkanes, ethers, alcohols,halogenated solvents, water, etc. Examples of the variety of solventsthat may be used include but are not limited to toluene, ethyl acetate,hexane, chlorobenzene, heptane, t-butyl methyl ether (MTBE), benzene,acetonitrile, cyclohexane, methylene chloride, 1,2-dichloroethane andt-butyl alcohol (t-BuOH), or a combination thereof. When usingRuW₁₁O₃₉SiNa₅ as the catalyst, heptane is the preferred solvent. WithRuCl₂(PPh₃)₃, chlorobenzene or benzene are preferred solvents.

This oxidation process may be run at a temperature between about −20° C.and up to the reflux temperature of the solvent used, for example about100° C., and particularly between about 5° C. and 50° C., and moreparticularly at about 15° C. The reaction may be run at any pH, andparticularly at an acidic pH, and more particularly at a pH of about 1.The pH of the reaction mixture may be adjusted prior to addition oft-BuOOH by addition of an aqueous acid such as sulfuric acid. Althoughnot required, the reaction is preferably run under an inert atmosphere,such as nitrogen or argon.

Δ-5-Steroidal alkenes that can be used in this process are known in theart. For example, see those listed and available through the SigmaChemical Co.

One embodiment of the present invention comprises the step of treating acompound of Formula I

with a hydroperoxide in the presence of a ruthenium based catalyst in asolvent to form a compound of Formula II

wherein Y is hydroxy, an esterified hydroxy group, keto or ethyleneketal, X is —CH₂—, —NH—, or —N(CH₃)— or —N-2,4-dimethoxybenzyl, and Z is

The oxidation reaction is not affected by the substituent at the 16- or17-position of the steroid, and thus “A” can be any syntheticallyfeasible substituent. The flexibility and broad applicability of theinstant process is demonstrated by the fact that it is not limited bythe choice of substituent at the 16- and 17-positions of the steroidalstarting material.

Representative examples of “A” include but are not limited to: —H; keto(═O); protected hydroxy, e.g. dimethyl-t-butyl silyloxy,trimethylsilyloxy, tri-ethylsilyloxy, tri-i-propylsilyloxy,triphenylsilyloxy; acetate; hydroxy; protected amino, e.g. acetylamino;amino; C₁₋₁₀ alkyl, e.g. methyl, ethyl, 1,5-dimethylhexyl,6-methylhept-2-yl cholestanyl 17-side chain, pregnane or stigmasterol17-side chain; aryl substituted C₁₋₁₀ alkyl, e.g. omega-phenylpropyl,1-(chlorophenoxy)ethyl; aryl carbamoyl substituted C₁₋₁₀ alkyl, e.g.2-(4-pyridinyl-carbamoyl)ethyl; C₁₋₁₀alkylcarbonyl, e.g.isobutylcarbonyl; arylcarbonyl, e.g. phenylcarbonyl; ether-substitutedC₁₋₁₀alkyl, e.g. 1-methoxy-ethyl, 1-ethoxy-ethyl; keto-substitutedC₁₋₁₀alkyl, e.g. 1-keto-ethyl; heteroaryl-substituted C₁₋₁₀ alkyl, e.g.omega-(4-pyridyl)-butyl; carboxy; carboxylic esters, e.g. C₁₋₁₀alkylcarboxylic esters such as carbomethoxy; carboxamides, e.g. C₁₋₁₀alkylcarboxamides or aralkylcarboxamides such as N,N-diisopropylcarboxamide, n-t-butyl carboxamide or N-(diphenylmethyl)-carboxamide;carbamates such as C₁₋₁₀ alkylcarbamates, especially t-butylcarbamate;substituted or unsubstituted anilide derivatives wherein the phenyl maybe substituted with 1 to 2 substitutents selected from ethyl, methyl,trifluoromethyl or halo (F, Cl, Br, I); ureas, e.g. C₁₋₁₀alkylcarbonylamino ureas such as t-butylcarbonylamino urea; C₁₋₁₀alkylcarbonylamino, e.g. t-butylcarbonylamino; ethers, e.g. n-butyloxy,ethylene ketal; substituted and unsubstituted aryl ethers such aschlorophenyloxy, methylphenyloxy, phenyloxy, methylsulfonylphenyloxy,pyrimidinyloxy; and the like.

The term “alkyl” includes both straight and branched chain alkyl groups,and “aryl” includes phenyl, pyridinyl and pyrimidinyl.

Hydroxy and amino protecting groups are known to those of ordinary skillin the art, and any such groups may be used. For example, acetate,benzoate, ether and silyl protecting groups are suitable hydroxyprotecting groups. Standard silyl protecting groups have the generalformula —Si(Xa)₃, wherein each Xa group is independently an alkyl oraryl group, and include, e.g. trimethylsilyl, tri-ethylsilyl,tri-i-propylsilyl, triphenylsilyl as well as t-butyl-di-(Xb)-silyl whereXb is methyl, ethyl, i-propyl or phenyl (Ph). Standard amino protectinggroups have the general formula —C(O)-Xc, wherein Xc is alkyl, aryl,O-alkyl or O-aryl, and include, e.g. N-t-butoxycarbonyl. See alsoProtective Groups in Organic Synthesis, T. W. Green et al. (John Wileyand Sons, 1991) for descriptions of protecting groups.

As will be appreciated by those of ordinary skill in the art, when Y isan esterified hydroxy group, substituents such as those of Formula IIIare intended

wherein Xd can form any synthetically feasible ester group. The processis not limited by the choice of any particular ester form for Y.Representative examples of Xa include but are not limited to straight orbranched chain alkyl, e.g. C₁₋₁₈ alkyl, phenyl, mono- or di-substitutedphenyl wherein the substituents include, e.g., halogen, alkoxy, andamino.

The intermediate compound II is useful for making 7β-substituted4-azasteroid compounds, and particularly those which are inhibitors of5α-reductase. Examples of such compounds include but are not limited tothose disclosed in U.S. Pat. Nos. 4,377,584 and 4,760,071; WO 93/23419;and WO 93/23420. More particularly, compounds that can be made fromintermediate II include those of general Formula IV:

wherein R is H or methyl, Z is

and Alk is selected from C₁₋₂₅ linear or branched alkyl, e.g., methyl(Me), ethyl (Et), propyl (Pr), isopropyl (i-Pr), n-butyl (n-Bu),sec-butyl, isobutyl, tert-butyl (t-Bu) and the like; C₃₋₆ cycloalkyl,e.g., cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl; and allyl.Processes for making such compounds are taught for example in U.S. Pat.No. 5,237,064, WO 93/23419 and WO 93/23420 and PCT application havingthe Ser. No. 94/12071.

A further exemplary synthetic scheme showing how to make compounds ofFormula IV is as follows:

The starting materials for the process generally are the3-acetoxy-androst-5-enes which are known and available in the art.

Z is

The term “A” is described above and can be any substituent preferablyinert and non-interfering under the particular reaction conditions ofeach step outlined in the above reaction scheme.

The A group can also be a protected hydroxy or protected amino groupwhich undergoes the indicated reaction sequence and then is subsequentlyremoved, or it can also be removed during a particular step providing itdoes not interfere with the indicated reaction. For example, where A is—O-TBDMS, i.e., t-butyldimethylsilyloxy, the silyl protecting group canbe removed during e.g., the ring closure step of the seco acid 6 to the4-aza steroid 7, such that the subsequent steps are performed on the 16-or 17-OH compound. Also, the starting A group can be a precursor to thefinally desired A group and be converted thereto concurrently in one ofthe steps. For example, where A contains a double bond, e.g., astigmasterol analog, the double bond in the 16- or 17-side chain mayalso be oxidized during the seco acid formation in going from 5 to 6.

As shown in the above Reaction Scheme, the “Alk” substituent can beintroduced onto the B ring of the 4-aza steroid generally by theapplication of an organometallic carbonyl addition reaction, e.g., theGrignard reaction in which the 7-carbonyl group can be reacted with theGrignard reagent containing “Alk” as the R radical in RMgX. The Grignardreaction conditions are conventional and include the use of, e.g.,methyl, allyl or cycloalkyl magnesium chloride, ethyl magnesium bromide,cyclopropyl magnesium bromide, and the like. Preferably, the Grignardreagent is used with CeCl₃. Usable dry solvents include, e.g.,tetrahydrofuran (THF), diethyl ether, dimethoxyethane, and di-n-butylether. The reaction is conducted under dry conditions generally in thetemperature range of 0° C. to 40° C. Generally, the reaction requiresabout 6 to 24 hours for completion. Other organometallic carbonyladdition reactions can be used in this step, such as those utilizinglithium and zinc organometallic reagents which are known in the art.

The adduct 3 is then oxidized with e.g. aluminum isopropoxide andcyclohexanone (Oppenauer oxidation conditions) in e.g. refluxing toluenesolvent to produce the 7-alkyl-4,6-dien-3-one 4. Other reagents whichcan be used are, e.g., aluminum ethoxide or aluminum t-butoxide. Othersolvents which can be used include, e.g., methylethylketone (MEK) andxylene. The temperature is generally in the range of about 60 to 120°C., and the reaction is carried out under anhydrous conditions andgenerally requires about 2 to 24 hours for completion.

The dien-3-one 4 is next converted to the 4-ene 5 by treatment with Pdon carbon, DBU, and cyclohexene in a solvent such as ethanol.

The A Ring is next cleaved by treatment with e.g. potassiumpermanganate, sodium periodate in e.g., t-butylalcohol at 80° C. toproduce the corresponding seco-acid 6. Other oxidation reagents whichcan be used include ruthenium tetraoxide and ozone. Other solvents whichcan be used are: CH₃CN, CCl₄, methanol (MeOH) and CH₂Cl₂. The reactiongenerally requires about 2 to 4 hours to proceed to completion.

The seco-acid in a C₂₋₄ alkanoic acid such as acetic acid (HOAc) istreated with ammonium acetate at about 15-30° C. followed by warming toreflux for about 2 to 4 hours. After cooling to about 50-70° C., wateris added and the mixture seeded to cause crystallization of theene-lactam 7.

Hydrogenation of the ene-lactam is accomplished with a noble metalcatalyst, such as a Pd(OH)₂, PtO₂, Pd on carbon, Rh on carbon orRh/Al₂O₃, and preferably using Rh on carbon or Rh/Al₂O₃, in a C₂₋₄alkanoic acid, such as acetic acid, or an alcohol such as ethanol, orethyl acetate, at about 50-70 psi hydrogen. The reaction is run at about15-25° C. for about 8 to 12 hours, and then the temperature may beraised, e.g., to about 50-70° C., until the reaction is essentiallycomplete. The catalyst is removed by filtration and the filtrate isconcentrated to dryness. The product 8 may then be purified, e.g., byrecrystallization.

The last step, N-methylation, is accomplished by treating a solution ofthe lactam 8 in an aromatic solvent such as benzene or toluene, in thepresence of tetrabutylammonium hydrogensulfate and aqueous alkali suchas potassium hydroxide or sodium hydroxide, with methyl chloride gaswith rapid stirring at about 40-60° C. until the reaction is essentiallycomplete, usually in about 20-30 hours.

Representative experimental procedures utilizing the novel process aredetailed below. These procedures are exemplary only and should not beconstrued as being limitations on the novel process of this invention.

EXAMPLE 1

Preparation of 4,7β-dimehyl-4-aza-5α-cholestan-3-one Step 1:

Materials Amt Mole MW Cholesteryl acetate (95% Aldrich) 78.1 gm 0.173428.7 t-BuOOH (70 wt %, Aldrich) 189 gm 1.46 90.12 Na₂WO₄-2H₂O 3.3 gm0.010 329.9 RUCl₃-xH₂O 0.24 gm 0.00116 207.43 Sodium metasilicate(Na₂SiO₃) 0.315 gm 0.00258 122.06 Sulfuric acid (d = 1.84 g/mL, 18 M)0.45 mL 0.0081 98.08 Sodium sulfite (Na₂SO₃) 39 gm 0.309 126.04 heptane300 mL MBK (methyl ethyl ketone) 550 mL water 460 mL

In a 2000 mL 3-necked flask was added sodium tungstate dihydrate (3.3gm), sodium metasilicate (0.315 gm) and 70 mL water and stirred untilhomogeneous. The solution was neutralized (pH=6-7) with concentratedsulfuric acid (0.45 mL). A 4° C. exotherm was noted for the addition ofacid. Ruthenium trichloride hydrate (240 mg) was added and the mixturestirred for 10 min. Cholesteryl acetate (78.1 gm) and heptane (300 mL)were added to the catalyst mixture. The stirring rate was 225-275 rpmwith an overhead paddle stirrer.

70% t-BuOOH (189 gm) was added over 5-10 min. An internal temperature of15-20° C. was maintained by cooling with a water bath. The temperatureof the batch began to rise slowly after an induction period of 5-15 min.The reaction was stirred until less than 1.5 wt % of s.m. (startingmaterial) and less than 2% of the 7-hydroxy cholesteryl acetateintermediate remained, about 20-24 hrs.

The reaction was monitored with a YMC basic column, 90:10acetonitrile:water, flow rate=1.5 mL/min, UV detection at 200 nm.Retention times: t_(R) cholesteryl acetate=17.0 min, t_(R) 7-ketocholesteryl acetate=7.8 min, t_(R) enedione 4.5 min, t_(R)7-hydroperoxides, 7-ols intermediates=6.8, 6.9, 7.0, 8.2 min. Latereluting impurities at 18 and 19 min are the 7-t-BuOO-cholesterylacetates.

To the reaction mixture was added 550 mL MEK, 390 mL water, and 39 gmssodium sulfite. The mixture was heated to 70° C. until the enedioneimpurity was gone, about 3 hrs. The reaction mixture cooled, then wastransferred to a separatory funnel and the aqueous layer cut and thenthe organic layer washed with 100 mL 1% brine. The MEK and t-BuOH werethen removed by an azeotropic distillation with heptane (800 mL heptaneadded after an initial concentration to 300 mL) until less than 0.7%combined MEK and t-BuOH remained as assayed by GC (gas chromatography).

The heptane was checked for MEK and tBuOH levels by GC using an HP-5column at 35° C. with a 0.5 mL flow rate. t_(R) MEK=4.9 min, t_(R)tBuOH=5.3 min, t_(R) heptane=7.7 min. The volume was adjusted to 350 mL,cooled to −5° C. and filtered, washing twice with 150 mL 0° C. heptane.After drying, the product was obtained in 62% yield (51.5 gms total, 94wt %, 97 A %) as an off-white solid. “A %” is area %.

Melting point (m.p.): 155-157° C.

NMR (¹H, 300 MHz, CDCl₃): 5.70 (s, 1H), 4.7 (m, 1H), 2.5-0.8 (m, 43 H),0.6 (s, 3H).

Step 2:

Materials Amt Mole MW 7-keto-cholesteryl acetate (95% pure) 60 gm (asis) 0.13 442 Methyl magnesium chloride (3.0 M) 160 mL 0.48CeCl_(3 (anhydrous)) 16.6 gm 0.068 245 THF (KF = 50 μg/mL) 300 mL Citricacid 115 gm 0.60 192 water 500 mL toluene 600 mL sat'd NaHCO₃ 240 mL

Anhydrous cerium chloride (16.6 gm) was stirred as a slurry in THF (150mL) at 20° C. under N₂ for 2 h.

The cerium chloride was obtained as the hepta-hydrate and dried in vacuoat an oven temperature of 170° C. for three to four days. The driedcerium chloride showed a weight loss of 0.7% by T.G. analysis. After twohours a sample of the slurry was removed and showed fine needles under amicroscope. To the slurry was added the Grignard reagent (160 mL) andthe resulting light purple mixture was aged for 30 minutes.

To the cooled mixture (20° C.) was added the ketone (60 gm at 95%purity, 57 gm by assay) in THF (150 mL) over 50 minutes while allowingthe mixture to exotherm to 30° C. Addition of the ketone to the Grignardreagent was exothermic, the exotherm was controlled by the rate ofaddition. The ketone solution in THF should be warmed to 30° C. toensure complete dissolution, prior to adding it to the Grignard reagent.

The reaction progress was monitored by HPLC (high pressure liquidchromatography). A 0.5 mL sample was added to 10 mL of 0.1N HOAc andthen diluted to 50 mL with CH₃CN. HPLC conditions [Zorbax® phenylcolumn, CH₃CN, water, phosphoric acid; 75:25:0.1 gradient elution to90:10:0.1 at 18 minutes, flow=1.5 mL/min, UV detection at 200 nm].Retention times, 3,7-diol t_(R)=5.6 and 5.9 min, starting ketonet_(R)=10.9 min, intermediate 7-OH, 3-OAc t_(R)=9.8 and 10.8 min. Therewas about 95 area % of 3,7 diol (ca. 85 mg/mL). (NOTE: Any remainingstarting material or reaction intermediates can be converted intoproduct using additional Grignard reagent.)

Once complete, the reaction was quenched by adding it to a 0° C. mixtureof citric acid solution (115 gm in 300 mL of water) and toluene (360mL). The quench was exothermic. (NOTE: The rate of addition should becarefully controlled to maintain an internal temperature below 10° C.)

The two phase mixture was stirred for 30 minutes and allowed to standfor 10-15 minutes for an adequate phase separation. The pH of theaqueous layer was ca. 2. The organic phase was separated, washed withwater (200 mL, pH=3 after washing) and saturated NaHCO₃ solution (240mL, pH=8 after washing). This afforded 750 mL of an organic layer whichcontained 66 mg/mL of diol for a yield of 49.5 gm (93%). The aqueouslayer contained less than 1% of product.

The batch was concentrated to 300 mL in vacuo (100-200 mm), diluted to600 mL with toluene and re-concentrated to 360 mL. The solvent switch totoluene was considered complete when the G.C. area % of THF was <2% ofthe toluene area %. (NOTE: The first 200 mL of the distillation has atendency to foam at low pressures. Once this phase is complete, thevacuum should be brought down to 100 mm. The distillation temperatureslowly rises from 20° C. to ca. 45° C. as the solvent switch to toluenenears completion.)

Samples of the distillate were assayed for residual THF using G.C. Asample of ca. 0.1 mL was diluted to 1 mL with methanol. G.C. conditions:[HP-5 column (25 M, 0.32 μm ID) using a heated block injector, 35° C.isothermal, flow=0.5 mL/min], MeOH t_(R)=5.5 min, THF t_(R)=6.2 min,toluene t_(R)=10.1 min. The final assay was performed using a samplefrom the batch.

The organic layer contained 134.4 mg/mL of diols for a total yield of48.4 gm (90%). (NOTE: The KF of the batch should be below 100 μg/mLbefore proceeding with the next step.)

Step 3: OPPENAUER OXIDATION

Materials Amt MMole MW 7-methyl-7-hydroxy-cholesterol 30.2 g 72.6 4162-butanone (d = 0.805, KF = 480 μg/mL) 126 mL 1404 72.11 Aluminumisopropoxide 18.9 g 93 204.25 3N HCl 120 mL 5% NaCl solution 120 mLConc. HCl 3.5 mL 42 D.I. water 60 mL Saturated NaHCO₃ 60 mL

To the toluene solution of the diol (256 mL, 118 mg/mL) was added2-butanone (126 mL) and aluminum isopropoxide (18.9 g). The solution washeated to reflux (92° C.) under nitrogen. The reaction progress wasmonitored by HPLC.

The batch was assayed for 2-butanone content by G.C. prior to adding thealuminum isopropoxide. A sample of ca. 0.1 mL was diluted to 1 mL withMeOH. G.C. conditions [HP-5 column (25 m, 0.32 μm ID) using a heatedblock injector at 250° C., column temp at 35° C. isothermal, flow=0.5mL/min] 2-butanone t_(R)=6.1 min, MeOH t_(R)=5.5 min, toluene t_(R)=10.1min. The KF of the starting mixture was 70 μg/mL.

A 0.1 mL sample of the reaction mixture was quenched into 0.1N HOAcsolution (2-3 mL) and then diluted to 10 mL with CH₃CN in a volumetricflask. HPLC conditions [25 cm Zorbax® Phenyl column; CH₃CN:H₂O with 0.1%phosphoric acid: 75:25 gradient elution to 90:10 at 18 min, hold 90:10until 22 min; flow=1.5 mL/min, UV detection at 210 nm.] Starting diolst_(R)=5.4, 5.8 min, intermediate Δ-4 eneone t_(R)=6.4 min, dieneonet_(R)=12.1 min.

The reaction was considered complete when the level of starting diol was<3 area % (8 hours). Once complete the batch was cooled to 15-20° C. andquenched with 3N HCl (120 mL). The two phase mixture was stirred for 20min, and then allowed to settle. The lower aqueous layer was removed andthe organic layer was washed with 5% NaCl (120 mL). The batch wasconcentrated in vacuo to one half volume (40-60° C. at 150 mm). Thedistillation removed excess 2-butanone from the batch. The level of2-butanone in the final batch was <2% of the toluene (using G.C.) andthe KF was 60 μg/mL.

The toluene solution was treated with conc. HCl (3.5 mL) at 25° C.,under N₂. The reaction was assayed by HPLC until the intermediatetertiary alcohol was completely converted to dieneone (ca. 1 h). Thesolution was washed with D.I. water (60 mL) and saturated NaHCO₃ (60mL). The pH of the bicarbonate wash was 8.5. (NOTE: The decompositionreaction will turn black if run for longer than 8 hours.) The resultingred solution (128 mL) contained 202 mg/mL of dienone for a yield of 25.9gm (90%).

Step 4: TRANSFER HYDROGENATION

Materials Amt MMole MW Dieneone (toluene solution) 31.5 g 79.5 396.7 5%Palladium on carbon (dry) 4.5 g Cyclohexene (d = 0.811) 120 mL 1.18 mole82.15 1,8 diazabicyclo[5.4.0]undec-7-ene (DBU) 0.63 mL 4.2 152.2Absolute ethanol 495 mL 3N HCl 150 mL half saturated NaHCO₃ 100 mL SolkaFlok Hexanes 250 mL t-butanol 175 mL

The toluene solution of the dieneone (150 mL at 214.6 mg/mL) was dilutedwith ethanol (120 mL) and cyclohexene (120 mL) and DBU (0.62 mL). To themixture was added 5% palladium on carbon (9.0 g of 50% water wet). Themixture was degassed using vacuum/nitrogen purges (3×). The slurry wasthen heated to reflux (reflux temperature=72° C.). The reaction wasmonitored by HPLC.

A 2 mL sample of the reaction mixture was filtered through Solka Floe.The filtrate (0.1 mL) was diluted to 10 mL with CH₃CN and analyzed byHPLC: 25 cm Zorbax® phenyl column; acetonitrile/water containing 0.1%phosphoric acid: gradient elution from 75:25 to 90:10 CH₃CN:water in 18min, hold 90:10 until 22 min; flow=1.5 mL/min; UV detection at 200 nm.

Dienone t_(R)=12.1 min, Δ-4 enone t_(R)=13.2 min, Δ-5 enone t_(R)=14.1min, over-reduced ketone t_(R)=14.4 min, ethyl enol ether t_(R)=20.9min. The over-reduced ketone should be assayed at 192 nm.

The reaction was considered complete when the dieneone level was <2 A %and the Δ-5 enone level was 5% (about 10 hours). When the reaction wascomplete the mixture was cooled to ambient temperature. The palladiumwas removed by filtration through Solka Floc and the filter cake waswashed with ethanol (150 mL).

The batch contained 51 mg/mL of enone. (NOTE: Prolonged reaction timesshould be avoided since over-reduction can occur. If the startingmaterial has been consumed and the level of Δ-5 enone is >5% after 10hours, then the palladium should be filtered, and the isomerizationcompleted without catalyst present.)

The solution was concentrated under reduced pressure (75 mm) to a volumeof approximately 150 mL. The batch was diluted with ethanol (225 mL) andre-concentrated to 150 mL.

The solvent switch to ethanol was considered complete when the toluenelevel was <2% of the ethanol by G.C., and there was no detectablecyclohexene. (NOTE: Removal of cyclohexene is important since it reactsin the subsequent oxidative cleavage step and unproductively consumesperiodate.) A 0.1 mL sample was diluted to 1 mL with ethanol for thecyclohexene assay (and 1,1,1 trichloroethane for the toluene assay).G.C. conditions [HP-5 (25M×0.32 μm ID), using a heated block injector at250° C., column temp at 35° C. isothermal, flow=0.5 mL/min] ethanolt_(R)=5.6 min, cyclohexene t_(R)=7.7 min, trichloroethane t_(R)=7.7 min,toluene t_(R)=10.2 min. The presence of cyclohexene is also detectableby ¹H NMR (CDCl₃) of the solution: cyclohexene vinyl protons at δ=5.64ppm, eneone vinyl proton at δ=5.69 ppm.

The concentrate was diluted with hexanes (250 mL) and 3N HCl (150 mL).The two phase mixture was warmed to 40° C. until enol ether hydrolysiswas complete. The layers were separated and the organic layer was washedwith half saturated sodium bicarbonate (100 mL). The hexane phase had avolume of 291 mL, contained less than 5% ethanol by volume and assayedfor 92 mg/mL of enone.

The solution was concentrated to 100 mL under reduced pressure (100 mm15° C.). The batch was diluted with t-butanol (175 mL) andre-concentrated to 100 mL (100 mm/40° C.). The batch contained 260 mg/mLof the desired 7-β-methyl enone for a yield of 26.8 gm (85%).

(NOTE: These compounds could also be detected by G.C.M.S. Use of G.C. tofollow this reaction should be avoided since the enone disproportionateson the column. G.C.M.S. conditions [HP-5 (25 M) column, on columninjection at 285° C. isothermal]; over-reduced enone t_(R)=12.8 min, 7alpha-epimer t_(R)=15.7 min, product t_(R)=17.3 min, s.m. t_(R)=21.3min.

Step 5: OXIDATIVE CLEAVAGE

Materials Amt Mol MW 7-β-Methylcholest-4-ene-3-one 300 gm 0.75 398t-Butanol (d = 0.786) 6.6 L Sodium carbonate 159 gm 1.5 106 Sodiumperiodate 1550 gm 7.2 213.9 Potassium permanganate 11.1 gm 0.07 158 D.I.Water 14.2 L Diatomite 50 gm Ethyl acetate (d = 0.902) 2.6 L Heptane (d= 0.684) 5.0 L conc. Hydrochloric acid 250 mL 5% Aqueous NaCl 2.5 LAcetic acid (d = 1.049) 9.0 L

In a 5 L roundbottom flask was charged D.I. water (4.93 L), sodiumperiodate (1.55 Kg) and potassium permanganate (11.1 gm). The slurry wasstirred at 65° C. for 30 minutes to obtain complete solution.

To a solution of the enone (300 gm) in t-butanol (4.60 L) was added asolution of sodium carbonate (159 gm) in water (2.3 L). The two phasemixture was warmed to 65° C. The enone should be toluene, ethanol andcyclohexene free. (NOTE: Concentration of enone in organic layer isabout 56 mgmL⁻¹.) The sodium periodate solution was added to the enonesolution over 3 h with rapid stirring, maintaining the reactiontemperature at 65° C. The slurry was aged at 65° C. for 2 h. Theperiodate solution was added via a heated addition funnel.

Carbon dioxide gas was evolved during the reaction. A slow additionensures controlled gas evolution. No exothermn was detected duringaddition. During the addition a purple/brown slurry was formed.

The reaction progress was monitored by HPLC. A 2 mL sample of thereaction mixture was cooled to 15° C. and filtered. The filtrate (0.1mL) was diluted to 10 mL with water/CH₃CN (1:3). HPLC conditions [YMCBasic 25 cm×4.6 mm, CH₃CN, 0.01M H₃PO₄; 90:10 isocratic flow=1.5 mL/min,UV detection at 200 nm]; enone t_(R)=11.5 min, seco-acid t_(R)=5.5 min.

The reaction was considered complete when the starting enone was <0.5mg/mL. Water (3.0 L) was added and the slurry heated to reflux for 2 hto decompose any remaining KMnO₄ (color change from purple to brown) andto dissolve most of the solids precipitated on the vessel walls. Theresultant slurry was cooled to 15° C. and filtered through dicalite (50gm). The vessel and cake were washed with t-butanol/water (1:2, 6.0 L).

The filter cake was assayed for seco acid by dissolving 200-400 mg ofcake with 50 mL water and 50 mL acetonitrile then filtering into thesample vial through diatomite to remove the small amount of orangemanganese solids. The filtrates (pH=9.0-10.5) were extracted withheptane (5.0 L).

Ethyl acetate (2.6 L) was added to the aqueous mixture and the pHadjusted to 2.5±0.3 by the addition of conc. HCl (250 mL). The aqueouslayer was removed.

The organic layer was washed with 5% aqueous brine (2×1.2 L). The ethylacetate solution was concentrated (150 mm.Hg, 30° C.) to approx. 10%volume. Acetic acid (7.4 L) was added and the residual ethyl acetateremoved by concentration (100 mm.Hg, 60° C.) to <1% by volume (<0.5 area% by HPLC). The final volume was adjusted to 5.0 L by addition of aceticacid. Ethyl acetate removal was monitored by HPLC using the conditionsabove except the flow rate was 0.5 mL min⁻¹ and UV detection at 210 nm.Ethyl acetate t_(R)=7.4 min, acetic acid t_(R)=6.9 min. The assay yieldwas 275 gm which represented an 88% yield. The acetic acid solution wasused directly in the following step (ene-lactam formation).

Step 6: NH-Enelactam Formation

Materials Amt Mole MW Seco-acid 265 gm 0.634 418 Ammonium Acetate 488 gm6.33 77.1 2,6-di-t-butyl-4- 5.3 gm 0.024 220 methylphenol (BHT) D.I.Water 565 mL Acetic acid 833 mL

To a solution of seco-acid in acetic acid (265 gm in 5.3 L) obtained inthe previous step was added BHT (5.3 gm) and ammonium acetate (488 gm)at 20° C. The slurry was warmed to a gentle reflux under a nitrogenatmosphere for 3 h. Complete solution was obtained at 30° C. Theinternal temperature was 120° C. at reflux. Color changed from yellow todark red/brown. Use of reduced amounts of acetic acid results in oilingof the product at the crystallization stage.

The reaction progress was monitored by UPLC. HPLC conditions [SB Phenyl,CH₃CN, 0.01M H₃PO₄; isocratic 80:20 for 30 min, flow=1.5 mL/min, UVdetection at 190/200 nm] Retention times: ene-lactam t_(R)=9.4 min, secoacid t_(R)=5.3 min. UV detection was at 190 nm for reaction progress and200 nm for s.m. and product assay. The reaction was considered completewhen <0.05 A % seco acid remained, about 3-4 hrs.

The reaction mixture was cooled to 60° C. and water (398 mL) added over15 min. (NOTE: Addition of exactly 7.5% v/v water to the acetic acidsolution is important.) The solution was allowed to cool to 50° C. andseeded with ene-lactam (1.0 gm). Crystallization occurred at 50° C. Theslurry obtained was aged at 50° C. for 1 h and then cooled to 0-2° C.over 2 h.

The slurry was filtered and the light tan solid washed with 5:1 aceticacid/water (1.0 L). The solid was dried in vacuo at 30° C. overnight togive 255 gm at 87 wt % by assay (remainder is acetic acid) for an 88%yield. HPLC profile, UV at 200 nm was 99.4 A %. Melting point (m.p.) ofsolvate=112-115° C. Pure m.p.=175-178° C., softens at 162° C.

Step 7: NH-Enelactam Recrystallization

Materials Amt Mole MW Enelactam 20 gm 0.041 400 D.I. Water 17 mL Aceticacid 133 mL BHT 0.20 gm 0.00091 220

To 20 gm at 83 wt % enelactam was added 100 mL acetic acid whichcontained 200 mg of BHT. The slurry was warmed to 60° C. under anitrogen atmosphere to achieve dissolution, then cooled to 50° C. Acharge of 10 mL water was then added. The mixture was then cooled to 5°C. over 1.5 hrs, aged for one hour and then the solid filtered off.(NOTE: The solution at 50° C. should have started crystallizing beforecooling to 5° C.) The solution KF after BHT addition was about 0.2-0.4%w/w.

The mother liquor amounts were monitored by HPLC. HPLC conditions [SBPhenyl, CH₃CN, 0.01M H₃PO₄, isocratic 80:20 for 30 min, flow=1.5 mL/min,UV detection at 200 nm] Retention times: ene-lactam t_(R)=9.4 min.Sample 100 μL and dilute to 10 mL with acetonitrile.

The slurry was filtered and the light tan solid washed with 5:1 aceticacid/water (40 mL) at 5° C. The solid was dried in vacuo at 30° C.overnight to give 18.5 gm at 84 wt % by assay (remainder is acetic acid)for a 94% recovery. HPLC profile, UV at 200 nm was 99.4 A %

M.p. solvate is 112-115° C. Pure m.p. is 175-178° C., softens at 162° C.

Step 8: N-H Enelactam Reduction

Materials Amount Mol MW Enelactam (87 wt %) 190.0 gm 0.475 399.64 HOAc(d = 1.05) 3.8 L BHT 3.8 gm 0.017 220.4 10% Pd/C (50% wet) 38 gm 10 wt %Hydrogen 60 psi Heptane 3.8 L MEK 2.65 L

BHT (3.8 gm) was dissolved in acetic acid (1.71 L) at 20° C. Thesolution was degassed with nitrogen purge for 30 min and the enelactam(218 gm at 87 wt %) added in one portion. The resultant solution waspurged with nitrogen for 15 minutes. 10% Pd/C (50% wet) (38 gm) wasadded and the slurry transferred to a 1 gallon stirred autoclave.Degassed acetic acid (190 mL) was used to rinse the slurry into theautoclave. (NOTE: BHT must be added to the acetic acid prior to additionof the ene-lactam. The use of BHT stabilized acetic acid is necessarydue to the oxidative instability of the ene-lactam.)

After vacuum purging with nitrogen the mixture was placed under 60 psiH₂ and stirred at 20° C. After 10 h at 20° C. the reaction temperaturewas increased to 60° C. until reaction was >99.9% complete.

The reaction was monitored by HPLC, [25.0 cm Zorbax® phenyl SB, 90:10CH₃CN: 0.01% H₃PO₄, 1.5 mL/min, Dual UV detection at 210 nm and 240 nm].Retention times: enelactam 8.50 min, trans-lactam 12.4 min, cis-lactam18.4 min. Sample 20 μL and dilute to 2 mL with acetonitrile.

On complete reaction (i.e., >99.9% conversion) the mixture was cooled to20° C. and filtered through Solka-Flok (20 gm). The cake was washed withacetic acid (1.9 L). The filtrates were combined and concentrated at 30°C./10 mm.Hg to a volume of 570 mL. Heptane (total of 3.8 L) was addedand concentration continued at atmospheric pressure (azeotropeb.p.=91-92° C.) to remove the acetic acid. (NOTE: Removal of the aceticacid to <0.2% by volume is important due to the very high solubility ofthe product in acetic acid.) Final b.p. was 98-99° C. Acetic acid wasmonitored at 200 nm by HPLC using a 25.0 cm Zorbax® phenyl SB columnwith 90:10 CH₃CN:water, 0.5 mL min⁻¹ as eluent. Sample 100 μL and diluteto 10 mL with acetonitrile.

The solution was concentrated to 570 mL and MEK (total of 2.5 L) added.The heptane was removed by azeotropic distillation at atmosphericpressure to <5% by volume as determined by G.C. of distillates andbatch. G.C. Conditions: DB-5 20 m. 0.5 mL min⁻¹ Helium, 35° C.isothermal; MEK t_(R)=6.4, heptane t_(R)=8.0 min. Crystallization occursduring removal of heptane.

The volume was adjusted to 600 mL and the solution was allowed to coolto 20° C. over 3 h. The resultant slurry was aged at −10° C. for 2 h.The solid was collected on a filter frit and washed with cold MEK (150mL). The solid was dried in vacuo at 20° C. Yield 170 gm, at >99 wt%; >99.2 A % at 210 nm. Step yield 89%.

Step 9: Methylation

Materials Amount Mol MW N-R Lactam 3.0 Kg 7.47 401.6 Methyl chloride 453gm 8.96 50.5 KOH/Alumina[1:1] 3.0 Kg 22.8 56 BnMe₃NCl 150 gm 0.81 185.7Toluene (d = 0.867) 14.0 L

A 5 gallon autoclave was charged with a slurry of lactam (3.0 Kg),BnMe₃NCl (150 gm) and potassium hydroxide on alumina (1:1, 3.0 Kg) intoluene (12 L) at room temperature. Methyl chloride (453 gm) wasintroduced at 20° C. with slow stirring. The slurry was heated to 65° C.with slow stirring and aged for 1 h. An exotherm at 52° C. of about 3°C. was noted as a spike on the temperature recorder.

The reaction progress was monitored by HPLC. HPLC conditions [Zorbax® SBphenyl, CH₃CN, 0.01M H₃PO₄; 90:10 isocratic, flow=1.5 mL/min, UVdetection at 200 nm] lactam t_(R)=12.4 min, IV-a t_(R)=15.0 min. 25 μLSample of toluene layer was diluted to 2 mL with acetonitrile. Thereaction was monitored until complete conversion was obtained (>99.95%).The reaction was complete in <60 min at 60° C.

The reaction mixture was cooled to 20° C. and purged with nitrogen (4×)to remove any excess MeCl. The toluene solution was filtered throughSolka Floc (100 gm) and the vessel and cake washed with toluene (2 L).The combined filtrates were concentrated at 100 mm. Hg and 20-30° C. toa residual oil. The oil should be homogeneous in heptane (10 mLg⁻¹)without any cloudiness.

The oil was assayed for toluene by G.C. oven temp 35° C. isothermal. Theproduct (100 mg) was dissolved in methanol (0.5 mL) and 1 μL injected.Toluene t_(R)=4.4 min, methanol t_(R)=2.7 min.

The oil was kept under vacuum until the solvent level was <2%. The oilwas poured into a glass tray and seeded with IV-a (1.25 gm) and allowedto stand in vacuo (20 mm.Hg) overnight.

The resulting solid was cut into blocks and broken up in a WARINGblender containing 2° C. water (10 L) to a particle size of <50 μm. Theslurry was filtered, washed with water (5.0 L) and dried in a nitrogenstream overnight. Yield of product=3.0 Kg, 97%.

EXAMPLE 2 Preparation of 7-keto cholesteryl acetate

The catalyst tris (triphenyl phosphine) ruthenium (II) chloride (96 mg,0.1 mmol) was dissolved in chlorobenzene (10 mL), followed bycholesteryl acetate (4.3 gm, 10.0 mmol). The mixture was degassed with avacuum nitrogen purge (3×), then cooled to +5° C. To the mixture underN₂ was added 90% t-butyl hydrogen peroxide (4.4 mL, 40 mmol) over 15hours. HPLC assay showed 2.85 gm (65%) of the 7-keto cholesterylacetate.

The batch was filtered through Solka-floc and the solvent was removed invacuo. The residue was dissolved in methanol and then the batch wascooled to +5° C. and aged for 30 minutes. The batch was filtered andwashed with cold methanol. The solid was air dried to afford 2.26 gm(51%) of 7-keto cholesteryl acetate.

Preparation of 7-keto cholesteryl acetate

Materials Amt Mole MW Cholesteryl acetate (95% Aldrich) 78.1 gm 0.173428.7 t-BuOOH (70 wt %, Aldrich) 229 gm 1.77 90.12 RUCl₃-xH₂O 0.24 gm0.00116 207.43 Sodium sulfite (Na₂SO₃) 39 gm 0.309 126.04 heptane 310 mLMEK (methyl ethyl ketone) 550 mL water 445 mL

In a 2000 mL 3-necked flask with an overhead stirrer was added rutheniumtrichloride hydrate (240 mg), 55 mL water, cholesteryl acetate (78.1 gm)and heptane (310 mL). The stirring rate was 225-275 rpm with an overheadpaddle stirrer.

70% t-BuOOH (229 gm) was added slowly over 4 hrs. An internaltemperature of 15-20° C. was maintained by cooling with a water bath.The temperature of the batch began to rise slowly after an inductionperiod of 5-15 min.

The reaction was stirred until less than 1.5 wt % of s.m. (startingmaterial) and less than 2% of the 7-hydroxy cholesteryl acetateintermediate remained, about 20-24 hrs.

The reaction was monitored with a YMC basic column, 90:10acetonitrile:water, flow rate=1.5 mL/min, UV detection at 200 nm.Retention times: t_(R) cholesteryl acetate=17.0 min, t_(R) 7-ketocholesteryl acetate=7.8 min, t_(R) enedione 4.5 min, t_(R)7-hydroperoxides, 7-ols intermediates=6.8, 6.9, 7.0, 8.2 min. Latereluting impurities at 18 and 19 min are the 7-tBuOO-cholesterylacetates.

To the reaction mixture was added 550 mL MEK, 390 mL water, and 39 gmssodium sulfite. The mixture was heated to 70° C. until the enedioneimpurity was gone, about 3 hrs. The reaction mixture was cooled thenfiltered through a pad of Solka-Flok to remove the ruthenium salts. Theclear solution was transferred to a separatory funnel and the aqueouslayer cut and then the organic layer washed with 100 mL 1% brine. TheMEK and t-BuOH were then removed by an azeotropic distillation withheptane (800 mL heptane added after an initial concentration to 300 mL)until less than 0.7% combined MEK and t-BuOH remained as assayed by GC(gas chromatography).

The heptane was checked for MEK and t-BuOH levels by GC using an HP-5column at 35° C. with a 0.5 mL flow rate. t_(R) MEK=4.9 min, t_(R)t-BuOH=5.3 min, t_(R) heptane=7.7 min.

The volume was adjusted to 350 mL, cooled to −5° C. and filtered,washing twice with 150 mL 0° C. heptane. After drying, the product wasobtained in 62% yield (51.5 gms total, 94 wt %, 97 A %), as an off-whitesolid.

EXAMPLE 4

Following essentially the same procedure as described in Example 1, Step1, the compounds of Formula II were made from the corresponding startingmaterials of Formula I wherein Z is

A, X and Y are defined as follows:

(a) A=6-methylhept-2-yl, X=—CH₂— and Y=—OH;

(b) A=ethylene ketal, X=—CH₂— and Y=ethylene ketal;

(c) A=t-butyl-di-methylsilyloxy (TBDMS-O—), X=—CH₂— and Y=—OC(O)CH_(3;)and

(d) A =6-methylhept-2-yl, X=—N(CH₃)— and Y=keto (═O)

Additionally, cyclohexenol was oxidized to cyclohexenone usingessentially the same procedure as described in Example 1, Step 1.

The starting material for (b) was prepared by treating commerciallyavailable 4-androstene-3,17-dione with ethylene glycol and HCl usingstandard reaction conditions. The starting material for (c) was preparedby treating commercially available 5-androsten-3,17-diol-3-acetate withTBDMS-Cl and imidazole using standard reaction conditions. The startingmaterial for (d) was prepared using standard synthetic procedures; i.e.,oxidative cleavage of commercially available cholestanone (proceduredescribed in Example 1, Step 5) followed by treatment with NH₂CH₃.

EXAMPLE 5

Materials Amt Mole MW 16-TBS Ether 100 gm* 0.224 447 Hexane 1000 mLRuthenium Chloride Hydrate 0.46 gm 0.0022 207.43 tBuOOH (70 wt %) 432 gm3.36 90.12 Sodium sulfite 20 gm 0.24 126 Water 600 mL Charcoal 10 gm *Wtassay by HPLC

To a solution of the 16-TBS ether (100 gm by assay, 16-tert.-butylsilylether) in hexane (500 mL) at 20° C. was added water (300 mL) andruthenium trichloride hydrate (0.46 gm). The two phase mixture wasstirred and cooled to 10° C. tBuOOH (70 wt %) (432 gm) was added over 5h while maintaining the reaction temperature 10-15° C.

The reaction is mildly exothermic. Water/ice was used to maintain thetemperature between 10 and 15° C.

The reaction was followed by HPLC employing a ZORBAX Phenyl SB, 25.0 cmcolumn, acetonitrile:water 30:70 to 80:20 over 25 min then held for 15min. UV detection at 200 nm. 1.5 mLmin⁻¹.

Retention times Min OTBS ether 29.4 7-Ketone 23.8 TBHP 3.25

The reaction is considered complete when <2% starting material remains(<1.5 mgmL-1).

Typical reaction time was 10 h.

After the reaction was completed, charcoal (10 gm) was added followed bysodium sulfite (20 gm) and the slurry stirred for 30 min.

The sodium sulfite decomposes any residual tBuOOH and otherhydroperoxides. Addition of sodium sulfite was mildly exothermic anddependent on the residual tBuOOH concentration. Complete removal oftBuOOH was checked by HPLC. The two phase mixture was filtered throughDICALITE (3″ in sintered funnel) and the cake washed with hexane (300mL). The aqueous layer was separated and the hexane layer washed withwater (100 mL×2).

A small rag layer at the interface was removed by addition ofacetonitrile (20 mL).

The hexane layer was concentrated to low volume and flushed with hexane(400 mL). The solution was concentrated to a final volume of 150 mL(approx. 2:1, hexane:substrate) and used ‘as is’ for the purificationstep.

The hexane solution was dried prior to silica treatment.

Silica Treatment

The hexane solution above was loaded onto a silica gel column (470 gm ,60-230 Mesh pre slurried in hexane). The column was eluted with hexane(800 mL) to remove unreacted starting material. The column was theneluted with 10% ethyl acetate in hexane (1000 mL) to provide the7-ketone.

Column Details 100 mL fractions, column was followed by TLC (20% ethylacetate/hex) or alternatively by HPLC (as above).

Fractions 14-17 were combined and concentrated to a total volume of100.0 mL.

While the foregoing specification teaches the principles of the presentinvention, with examples provided for the purpose of illustration, itwill be understood that the practice of the invention encompasses all ofthe usual variations, adaptations, and modifications, as come within thescope of the following claims and its equivalents.

What is claimed is:
 1. A process for oxidizing a Δ-5-steroidal alkene tothe corresponding Δ-5-7-keto-steroidal alkene comprising treating theΔ-5-steroidal alkene in solvent with a hydroperoxide in the presence ofa ruthenium-based catalyst.
 2. The process of claim 1 wherein thetemperature of the reaction is from about −20° C. to about 100° C. 3.The process of claim 2 wherein the temperature is from about 5° C. toabout 50° C.
 4. The process of claim 3 wherein the temperature is about15° C.
 5. The process of claim 1 wherein the reaction is ran at anacidic pH.
 6. The process of claim 5 wherein the pH is about
 1. 7. Theprocess of claim 1 wherein the reaction is run under an inertatmosphere.
 8. The process of claim 1 wherein the solvent is selectedfrom: water, toluene, ethyl acetate, hexane, chlorobenzene,1,2-dichloroethane, heptane, t-butyl methyl ether, benzene,acetonitrile, cyclohexane, methylene chloride, t-butyl alcohol, andmixtures thereof.
 9. The process of claim 1 wherein the ruthenium-basedcatalyst is a ruthenium sodium tungstate-based catalyst.
 10. The processof claim 9 wherein the ruthenium sodium tungstate-based catalyst isRuW₁₁O₃₉SiNa₅.
 11. The process of claim 1 wherein the ruthenium-basedcatalyst is selected from RuW₁₁O₃₉SiNa₅, RuCl₃, RuCl₂(PPh₃)₃, Ru(acac)₃,Ru(dimethylglyoximato)₂(PPh₃)₂, RuO₂, Ru/C, Ru(TPP)(CO)(THF),Ru(bipy)₂Cl₂, and K₅SiRu(H₂O)W₁₁O₃₉.
 12. The process of claim 1 whereinthe hydroperoxide is selected from t-butyl hydrogen peroxide, cumenehydroperoxide, hydrogen peroxide, and benzoyl peroxide.
 13. The processof claim 12 wherein the hydroperoxide is t-butyl hydrogen peroxide. 14.The process of claim 13 wherein the ruthenium-based catalyst isRuW₁₁O₃₉SiNa₅.
 15. The process of claim 14 wherein the solvent isheptane.
 16. The process of claim 1 wherein the Δ-5-steroidal alkene andthe Δ-5-7-keto-steroidal alkene have Formulas I and II, respectively:

wherein Z is

Y is selected from hydroxy, an esterified hydroxy group, keto andethylene ketal; X is selected from —CH₂—, —NH— or —N(CH₃)— or—N-2,4-dimethoxybenzyl; and A is selected from: —H, keto, protectedhydroxy, acetate, hydroxy, protected amino, amino, C₁₋₁₀ alkyl,aryl-substituted C₁₋₁₀ alkyl, aryl carbamoyl-substitutedC₁₋₁₀alkylC₁₋₁₀alkylcarbonyl, arylcarbonyl, ether-substitutedC₁₋₁₀alkyl, keto-substituted C₁₋₁₀alkyl, heteroaryl-substituted C₁₋₁₀alkyl, carboxylic ester, carboxamide, carbamate, substituted andunsubstituted anilide derivatives, urea, C₁₋₁₀ alkylcarbonylamino,ethylene ketal, ether and substituted and unsubstituted aryl ether. 17.The process of claim 16 wherein (a) protected hydroxy is selected fromdimethyl-t-butyl silyloxy, trimethylsilyloxy, tri-ethylsilyloxy,tri-i-propylsilyloxy, and triphenylsilyloxy; (b) protected amino isacetylamino; (c) C₁₋₁₀ alkyl is selected from methyl, ethyl,1,5-dimethylhexyl, 6-methylhept-2-yl, and 1-methyl-4-isopropylhexyl; (d)aryl substituted C₁₋₁₀ alkyl is selected from omega-phenylpropyl, and1-(chlorophenoxy)ethyl; (e) aryl carbamoyl substituted C₁₋₁₀ alkyl is2-(4-pyridinyl-carbamoyl)ethyl; (f) C₁₋₁₀alkylcarbonyl isisobutylcarbonyl; (g) arylcarbonyl is phenylcarbonyl; (h)ether-substituted C₁₋₁₀alkyl is selected from 1-methoxy-ethyl and1-ethoxy-ethyl; (i) keto-substituted C₁₋₁₀alkyl is 1-keto-ethyl; (j)heteroaryl-substituted C₁₋₁₀ alkyl is omega-(4-pyridyl)-butyl; (k)carboxylic esters are C₁₋₁₀ alkylcarboxylic esters selected fromcarbomethoxy and carboethoxy; (l) carboxamides are selected fromN,N-diisopropyl carboxamide, N-t-butyl carboxamide andN-(diphenylmethyl)-carboxamide; (m) carbamates are selected fromt-butylcarbamate and isopropylcarbamate; (n) substituted orunsubstituted anilide derivatives are selected from wherein the phenylmay be substituted with 1 to 2 substitutents selected from ethyl,methyl, trifluoromethyl or halo (F, Cl, Br, I); (o) urea ist-butylcarbonylamino urea; (p) C₁₋₁₀ alkylcarbonylamino ist-butylcarbonylamino; (q) ether is selected from n-butyloxy and ethyleneketal; (r) substituted and unsubstituted aryl ether is selected fromchlorophenyloxy, methylphenyloxy, phenyloxy, methylsulfonylphenyloxy andpyrimidinyloxy.
 18. The process of claim 16 wherein Z is


19. The process of claim 18 wherein A is selected from:6-methylhept-2-yl, t-butylcarbamoyl, phenylcarbamoyl,2,5-ditrifluoromethylphenylcarbamoyl, 4-methylsulfonyl-phenoxy,isobutylcarbonyl, phenylcarbonyl, 1-methoxyethyl, 1-keto-ethyl,2-(4-pyridinylcarbamoyl)ethyl, and chlorophenoxyethyl.
 20. The processof claim 18 wherein A is 6-methylhept-2-yl.
 21. The process of claim 16wherein Z is


22. The process of claim 21 wherein A is: phenoxy, chlorophenoxy,methylphenoxy, 2-pyrimidinyloxy, and tert.-butylsilyloxy.
 23. Theprocess of claim 16 wherein the ruthenium-based catalyst is a rutheniumsodium tungstate-based catalyst.
 24. The process of claim 23 wherein theruthenium sodium tungstate-based catalyst is RuW₁₁O₃₉SiNa₅.
 25. Theprocess of claim 24 wherein Y is —OC(O)CH₃ and X is —CH₂—.